Mercury is a highly toxic heavy metal in the environment. Mercury exposure can cause a number of severe adverse health effects, such as damages in the brain, nerve system, immune system, kidney, and many other organs.[1] Mercury contamination comes from both nature and human activities, and an annual releasing of 4,400 to 7,500 metric tons of mercury to the environment was estimated by the United Nations Environment Programme (UNEP).[2] Therefore, highly sensitive and selective mercury sensors are very useful in understanding its distribution and pollution and in preventing mercury poisoning. Towards this goal, many fluorescent small organic molecule-based Hg2+ sensors have been reported, which change their emission properties upon binding to Hg2+. Most of these sensors, however, require the involvement of organic solvent, show quenched emissions, and suffer from poor selectivity.[3-11] Only a few such sensors can detection Hg2+ in water with high sensitivity and selectivity.[12-17] Hg2+ sensors based on foldamers,[18, 19] oligonucleotides,[20] genetically engineered cells,[21] enzymes,[22] antibodies,[23] transcriptional regulatory proteins,[24,25] DNAzymes,[26] and chemically modified optical fibers[27,28] capillary optode,[29,30] membranes,[31] electrodes,[32,33] mesoporous silica,[34] and nanoparticles[35] are also known. For environmental monitoring applications, such as detection of Hg2+ in drinking water, a detection limit of lower than 10 nM (the toxic level defined by the US Environmental Protection Agency (EPA)) is required. However, few reported mercury sensors can reach such sensitivity.[11,21,25] We are interested in using catalytic DNA or DNAzymes to design metal sensors that can achieve the goal.[36,37]
DNAzymes are DNA-based biocatalysts.[38-42] Similar to protein enzymes or ribozymes, DNAzymes can also catalyze many chemical and biological transformations, and some of the reactions require specific metal ions as cofactors. Highly effective fluorescent and colorimetric sensors have been demonstrated for Pb2+ and UO22+ with DNAzymes.[36,37,43] These sensors showed picomolar to low nanomolar sensitivity and thousand to million-fold selectivity. In the presence of target metal ions, the fluorescence enhancement was generally greater than 10-fold, and signal generation took only 2 min or less. These sensors can be used at room temperature in aqueous solutions and no organic solvents are needed. Recently, DNAzyme-based electrochemical metal sensors are also reported.[44] Compared to protein or RNA, DNA is relatively more cost-effective to produce and more stable. DNAzymes can be denatured and renatured many times without losing their activities.[39] Therefore, DNAzymes are useful in metal detection.
It was reported that Hg2+ can specifically bind in between two DNA thymine bases and promote such T-T mismatches into stable base pairs (FIG. 1d).[20,45,46] This property was applied by Ono and co-workers to design a fluorescent sensor for Hg2+ detection.[20] The sensor consisted of a single-stranded thymine rich DNA with the two ends labeled with a fluorophore and a quencher, respectively. In the presence of Hg2+, the two ends were brought close to each other, resulting in decreased fluorescence. A detection limit of 40 nM was reported.[20] Being sensitive and selective, this sensor is a “turn-off” sensor and fluorescence intensity decreased in the presence of Hg2+, which may give false positive results caused by external quenchers or other environmental factors that can also induce fluorescence decrease. The Hg2+ stabilization effects on T-T mismatches have also been applied to design colorimetric sensors with DNA-functionalized gold nanoparticles and a detection limit of 100 nM was achieved.[35] In previous DNAzyme work, a signaling method called catalytic beacon was designed in which the metal binding site in DNAzymes and the fluorescence signaling part are spatially separated.[36,37,47]